Microfluidic test module with low mass electrochemiluminescent probe spots

Abstract

A microfluidic test module for detecting target nucleic acid sequences in a fluid, the test module having an outer casing having an inlet for receiving the fluid containing the target nucleic acid sequences, electrode pairs for receiving an electrical pulse, electrochemiluminescent (ECL) probe spots in contact with each of the electrode pairs respectively, the ECL probe spots containing ECL probes having an ECL luminophore for emitting photons when in an excited state and a functional moiety for quenching photon emission from the ECL luminophore by resonant energy transfer, such that the electrical pulse to the electrode pair excites the ECL luminophores, wherein, the mass of the ECL probes in each of the probe spots is less than 270 picograms.

1. A microfluidic test module for detecting target nucleic acid sequences in a fluid, the test module comprising:
 an outer casing having an inlet for receiving the fluid containing the target nucleic acid sequences; 
 electrode pairs for receiving an electrical pulse; 
 electrochemiluminescent (ECL) probe spots in contact with each of the electrode pairs respectively, the ECL probe spots containing ECL probes having an ECL luminophore for emitting photons when in an excited state and a functional moiety for quenching photon emission from the ECL luminophore by resonant energy transfer, such that the electrical pulse to the electrode pair excites the ECL luminophores; 
 an array of hybridization chambers containing the ECL probes for different target nucleic acid sequences; 
 a plurality of calibration sources each configured to provide a calibration emission, the calibration sources being distributed throughout the array of hybridization chambers such that each of the calibration sources are surrounded by a three-by-three square of the hybridization chambers; wherein, 
 the mass of the ECL probes in each of the probe spots is less than 270 picograms. 
 
     
     
       2. The microfluidic device according to  claim 1  wherein the mass of the probes in each of the probe spots is less than 60 picograms. 
     
     
       3. The microfluidic device according to  claim 2  wherein the mass of the probes in each of the probe spots is less than 12 picograms. 
     
     
       4. The microfluidic device according to  claim 3  wherein the mass of the probes in each of the probe spots is less than 2.7 picograms. 
     
     
       5. The microfluidic test module according to  claim 1  wherein the ECL luminophore has a transition metal-ligand complex. 
     
     
       6. A microfluidic test module according to  claim 1  further comprising:
 a detection photosensor for exposure to the photons emitted by the ECL luminophores; and, 
 control circuitry for providing the electrical pulse to the electrodes. 
 
     
     
       7. A microfluidic test module according to  claim 6  further comprising:
 a communication interface for the control circuitry to transmit data to an external device. 
 
     
     
       8. A microfluidic test module according to  claim 7  wherein the control circuitry has memory for storing the identity data relating to the ECL probes in each of the hybridization chambers. 
     
     
       9. A microfluidic test module according to  claim 8  wherein the hybridization chambers have a volume less than 900,000 cubic microns. 
     
     
       10. A microfluidic test module according to  claim 9  wherein the hybridization chambers have a volume less than 200,000 cubic microns. 
     
     
       11. A microfluidic test module according to  claim 7  wherein the communication interface is a universal serial bus (USB) connection such that the outer casing is configured as a USB drive. 
     
     
       12. A microfluidic test module according to  claim 8  wherein the detection photosensor is an array of detection photodiodes positioned in registration with the hybridization chambers. 
     
     
       13. The microfluidic test module according to  claim 12  further comprising:
 a calibration photosensor for sensing the calibration emission wherein the control circuitry has a differential circuit for subtracting the calibration photosensor output from each of the detection photodiode outputs. 
 
     
     
       14. The microfluidic test module according to  claim 13  wherein the calibration photosensor is a plurality of the calibration photodiodes in registration with the calibration sources respectively. 
     
     
       15. The microfluidic test module according to  claim 14  wherein the calibration sources are calibration probes without an ECL luminophore. 
     
     
       16. The microfluidic test module according to  claim 14  further comprising a plurality of calibration chambers containing the calibration sources distributed throughout the array of hybridization chambers, wherein during use, output from any one of the detection photodiodes is compared to output from the calibration photodiode most proximate to that detection photodiode. 
     
     
       17. The microfluidic test module according to  claim 16  wherein the calibration sources are calibration probes and the calibration chambers are configured to seal the calibration probes from the fluid containing the target nucleic acid sequences. 
     
     
       18. The microfluidic test module according to  claim 12  wherein the detection photodiodes are less than 1600 microns from the hybridization chambers. 
     
     
       19. The microfluidic test module according to  claim 1  wherein the ECL probes have a stem-and-loop structure with a loop portion containing the sequence complementary to the target nucleic acid sequence, the loop portion being positioned between the functional moiety for quenching photon emission from the ECL luminophore, and the ECL luminophore, such that hybridization with the target nucleic acid sequence opens the loop portion and moves the ECL luminophore away from the functional moiety.